Clinical observations and experimental animal models implicate the spleen as an important site for host immune responses to blood-borne bacteria. How the spleen's unique tissue structure and the diverse mixture of resident myeloid and lymphoid cells work together in vivo to combat infection remains poorly understood. We expect the native tissue environment to regulate immune cell function in complex and profound ways and therefore in vivo imaging approaches are a crucial complement biochemical and in vitro studies. Using two-photon microscopy and a robust histological analysis, we propose to investigate early phagocyte clearance mechanisms, analyze the trafficking of splenic phagocytes and the role of chemokine signaling in spleen remodeling, and identify the antigen presenting cells (APCs) and tissue microenvironment involved in presenting bacteria-specific antigen to T cells. Our overall goal is to understand how spleen structure impacts immune function. Our hypothesis is that early host-pathogen interactions in the marginal zone induce bactericidal responses in some cells, while in others, those interactions trigger cell migration to the appropriate tissue microcompartments for antigen presentation. Our studies will use the well characterized Listeria model of bacterial infection, in which splenic infection leads to T cell priming and induces long-lasting immunity. The proposed studies will enhance our fundamental understanding of how the spleen responds to blood-borne bacteria and provide a framework for future studies of other more virulent human pathogens.